The present invention relates to the use of papillomavirus L2 protein in medicine, particularly for the regression of papillomavirus tumors in mammals; and to pharmaceutical formulations comprising the L2 protein.
Papillomaviruses induce a variety of lesions both in humans and in animals. Some papillomas, albeit benign, are themselves a clinical problem, such as laryngeal papillomas of children (Steinberg and Abramson, 1985) or penile papillomas of bulls (Jarrett, 1985a), and others are known to be a risk factor in the pathogenesis of cancer, as in the case of flat lesions of the cervix or penile condylomata in humans (zur Hausen, 1978). Therefore both in human and veterinary medicine an antiviral vaccine, particularly a therapeutic one inducing lesion rejection, would be of major importance. Vaccination studies in humans present several problems: first of all experimentation is ethically unacceptable and, secondly, very limited amounts of virus are available as some lesions, in particular those of the cervix, do not produce viral progeny, and no in vitro system is yet available which allows vegetative replication of virus. The production of viral proteins in bacteria and the use of synthetic peptides have circumvented this last problem and have allowed the ongoing analysis of the immune response to papillomavirus infection (see for instance Jenison et al, 1988: Jochmus-Kudielka et al, 1989; Tindle et al, 1990, Dillner, 1990 and Strang et al, 1990). Whilst investigation into the feasibility of a human papillomavirus vaccine is still at an early stage, effective prophylactic vaccines, both natural (Jarrett et al, 1990a) and genetically engineered (Pilachinski et al, 1986) have already been produced against bovine papillomaviruses, and regression of Shope papillomas has been achieved by vaccinating rabbits with tumour tissue extracts (Evans et al, 1962). The bovine system is an excellent model for the human one, given the several similarities between the two: multiple virus types with high lesion specificity (Campo et al, 1981; Jarrett et al, 1984), homology of genetic structure (Danos et al, 1984) and progression of some lesions to malignancy (Jarrett et al, 1978). The bovine system also presents several advantages: cofactors in oncogenesis are known (Jarrett et al, 1978; Campo and Jarrett, 1986) and, above all, direct experimentation is possible (Jarrett, 1985a).
It has recently been shown that vaccination with bovine papillomavirus type 2 (BPV-2) successfully prevented infection by the same virus (Jarrett et al, 1990a), but not by other virus types (Jarrett et al, 1990b). Prevention was accompanied by production of neutralising antibodies in the serum of vaccinated animals, indicating that neutralising epitopes are present in the virus.
Generally speaking, the present invention resides in the discovery that the papillomavirus L2 protein is therapeutically effective in the treatment of papillomavirus tumors or lesions.
Thus, the present invention provides the use of papillomavirus L2 protein in medicine, particularly for therapy of papillomavirus tumors or lesions.
The invention also provides a pharmaceutical formulation for the therapy of papillomavirus tumors or lesions, which comprise papillomavirus L2 protein in admixture with a pharmaceutically acceptable carrier.
The invention further provides papillomavirus L2 protein for use in the production of a medicament for use in medicine, particularly for use in the therapy of papillomavirus tumors or lesions.
The invention still further provides a method of treating a mammal for the therapy of papillomavirus tumors or lesions, which comprise the administration of papillomavirus L2 protein to the mammal.
Generally speaking, the therapeutic effect of the L2 protein may be limited to the respective papillomavirus type. Thus, for general therapeutic applications, especially where the particular papillomavirus type is unknown, it may be desirable to employ a mixture of L2 proteins from a variety of papillomavirus types.
Generally, the therapy will be applicable to papillomavirus infections of mammals, including humans and animals. In humans, the invention is particularly applicable for the therapy and regression of laryngeal tumors, skin cancer tumors and genital lesions, whether malignant or not. In animals, the therapy is particularly useful for the regression of tumors on animals, for example the removal of warts from the udders of milk cows, or removal of papillomas of the alimentary canal and for the treatment of horses and donkeys. Prophylactic vaccination may also be employed.
The L2 protein is generally produced by recombinant DNA techniques. In particular, a plasmid containing the gene coding for the L2 protein may be transfected into E. coli and cultured. The entire L2 protein as it exists in nature may be employed, or a fragment (such as amino acid 90 to 467 of BPV-2 as disclosed hereafter) or fragments thereof may be used providing that the therapeutic effectiveness is retained. The L2 protein may be the native form, with additions, deletions or substitutions which do not substantially effect its therapeutic effectiveness.
The L2 protein will usually be administered in the form of a pharmaceutical formulation. The formulation contains a pharmaceutically acceptable carrier. The carrier must be acceptable in the sense of being compatible with the other ingredients of the formulation and not deleterious to the recipient thereof.
Since the protein is broken down in the stomach, oral administration is not preferred. The pharmaceutical formulation is preferably formulated for parenteral administration, including subcutaneous, intramuscular and intravenous injection; or as a suppository or pessary. For parenteral administration the formulation may be presented as a sterile solution or suspension in a suitable liquid vehicle, which may also contain preservatives and materials for rendering the formulation isotonic. The formulations may be presented in unit-dose or multi-dose containers. The carrier will generally be apyrogenic. Each dose will generally contain 100 to 10,000 micro grams of the L2 protein.
In order to enhance the therapeutic effect of the protein, it may be administered together with an adjuvant, such as Freund""s incomplete adjuvant, as an oil-in-water emulsion or using other adjuvant systems known in the art such as L101 and DDA as used in Pilacinski et al. (1986).